Basil variety ‘eleonora’

ABSTRACT

New basil variety designated ‘Eleonora’ is described. ‘Eleonora’ is a basil variety exhibiting stability and uniformity.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of U.S. Provisional Application No.61/936,101, filed Feb. 5, 2014, which is hereby incorporated byreference in its entirety.

FIELD OF THE INVENTION

The present invention relates to the field of plant breeding. Inparticular, this invention relates to new basil, Ocimum basilicumvarieties designated ‘Elidia’, ‘Keira’, and ‘Eleonora’.

BACKGROUND OF THE INVENTION

Basil, Ocimum basilicum, is a popular herb having cooking and medicinaluses. Ocimum basilicum is in the Lamiaceae (mint) family. Basil is alsoknown as Thai basil, sweet basil, and Saint Joseph's Wort. Basil isnative to India and other tropical regions of Asia. There are manyvarieties of basil, as well as several related species or specieshybrids also called basil. The type used in Italian food is generallyknown as sweet basil, while in Asia common basil varieties are known asThai basil (O. basilicum var. thyrsiflora), lemon basil (O. Xcitriodorum) and holy basil (Ocimum tenuiflorum). Most common varietiesof basil are treated as annuals; however, some basil varieties areperennial in warm, tropical climates.

In cooking, basil is commonly used either fresh or dried to impart itsdistinctive flavor into various dishes, especially in Italian cuisine.The most common types of basil for cooking purposes are the SweetItalian basil varieties. As an herbal medicine, basil is believed tohave a soothing effect on the digestive system. Recently, the demand forfresh basil has mushroomed. Not only has there been a general trend incooking to use fresh ingredients, but modern cooks are discovering thetaste advantages of using fresh herbs such as basil.

Basil is an important and valuable herb. Accordingly, there is a needfor new basil varieties. In particular, there is a need for improvedflat basil varieties that are stable, high yielding, and agronomicallysound.

SUMMARY OF THE INVENTION

In order to meet these needs, the present invention is directed toimproved basil varieties.

As used herein basil variety ‘Elidia’ is the same basil variety as basilvariety ‘E9311732’ having ATCC Accession Number X1 and disclosed in U.S.Provisional Application No. 61/936,101. While the name has changed,basil variety ‘Elidia’ has all the defining characteristics of basilvariety ‘E9311732’.

As used herein basil variety ‘Keira’ is the same basil variety as basilvariety ‘E09B1002’ having ATCC Accession Number X2 and disclosed in U.S.Provisional Application No. 61/936,101. While the name has changed,basil variety ‘Keira’ has all the defining characteristics of basilvariety ‘E09B1002’.

As used herein basil variety ‘Eleonora’ is the same basil variety asbasil variety ‘E9311540’ having ATCC Accession Number X3 and disclosedin U.S. Provisional Application No. 61/936,101. While the name haschanged, basil variety ‘Eleonora’ has all the defining characteristicsof basil variety ‘E9311540’.

In one embodiment, the present invention is directed to basil, Ocimumbasilicum, seed designated as ‘Elidia’. In one embodiment, the presentinvention is directed to an Ocimum basilicum basil plant and partsisolated therefrom produced by growing ‘Elidia’ basil seed. In anotherembodiment, the present invention is directed to an Ocimum basilicumplant and parts isolated therefrom having all the physiological andmorphological characteristics of a Lactuca sativa plant produced bygrowing ‘Elidia’ basil seed. In still another embodiment, the presentinvention is directed to an F₁ hybrid Ocimum basilicum basil seed,plants grown from the seed, and a head isolated therefrom having‘Elidia’ as a parent, where ‘Elidia’ is grown from ‘Elidia’ basil seed.

Basil plant parts include basil stems, basil leaves, parts of basilleaves, pollen, ovules, flowers, roots, cells, and the like. In anotherembodiment, the present invention is further directed to basil stems,basil leaves, parts of basil leaves, flowers, pollen, and ovules, roots,or cells isolated from ‘Elidia’ basil plants. In another embodiment, thepresent invention is further directed to tissue culture of ‘Elidia’basil plants, and to basil plants regenerated from the tissue culture,where the plant has all of the morphological and physiologicalcharacteristics of ‘Elidia’ basil plants.

In still another embodiment, the present invention is further directedto packaging material containing ‘Elidia’ plant parts. Such packagingmaterial includes but is not limited to boxes, plastic bags, etc. The‘Elidia’ plant parts may be combined with other plant parts of otherplant varieties.

In yet another embodiment, the present invention is further directed toa method of selecting basil plants, by a) growing ‘Elidia’ basil plantswhere the ‘Elidia’ plants are grown from basil seed and b) selecting aplant from step a). In another embodiment, the present invention isfurther directed to basil plants, plant parts and seeds produced by thebasil plants where the basil plants are isolated by the selection methodof the invention. In another embodiment, the present invention isfurther directed to a method of breeding basil plants by crossing abasil plant with a plant grown from ‘Elidia’ basil. In still anotherembodiment, the present invention is further directed to basil plants,basil parts from the basil plants, and seeds produced therefrom wherethe basil plant is isolated by the breeding method of the invention. Insome embodiments, the basil plant isolated by the breeding method is atransgenic basil plant. In another embodiment, the present invention isdirected to methods for producing a basil plant containing in itsgenetic material one or more transgenes and to the transgenic basilplant produced by those methods. In another embodiment, the presentinvention is directed to methods for producing a male sterile basilplant by introducing a nucleic acid molecule that confers male sterilityinto a basil plant produced by growing ‘Elidia’ basil seed, and to malesterile basil plants produced by such methods.

In another embodiment, the present invention is directed to methods ofproducing an herbicide resistant basil plant by introducing a geneconferring herbicide resistance into a basil plant produced by growing‘Elidia’ basil seed, where the gene is selected from glyphosate,sulfonylurea, imidazolinone, dicamba, glufosinate, phenoxy proprionicacid, L-phosphinothricin, cyclohexone, cyclohexanedione, triazine, andbenzonitrile. Certain embodiments are also directed to herbicideresistant basil plants produced by such methods. In another embodiment,the present invention is directed to methods of producing a pest orinsect resistant basil plan by introducing a gene conferring pest orinsect resistance into a basil plant produced by growing ‘Elidia’ basilseed, and to pest or insect resistant basil plants produced by suchmethods. In certain embodiments, the gene conferring pest or insectresistance encodes a Bacillus thuringiensis endotoxin. In anotherembodiment, the present invention is directed to methods of producing adisease resistant basil plant by introducing a gene conferring diseaseresistance into a basil plant produced by growing ‘Elidia’ basil seed,and to disease resistant basil plants produced by such methods. Inanother embodiment, the present invention is directed to methods ofproducing a basil plant with a value-added trait by introducing a geneconferring a value-added trait into a basil plant produced by growing‘Elidia’ basil seed, where the gene encodes a protein selected from aferritin, a nitrate reductase, and a monellin. Certain embodiments arealso directed to basil plants having a value-added trait produced bysuch methods.

In another embodiment, the present invention is directed to methods ofintroducing a desired trait into basil variety by: (a) crossing a‘Elidia’ plant, with a plant of another basil variety that contains adesired trait to produce progeny plants, where the desired trait isselected from male sterility; herbicide resistance; insect or pestresistance; modified bolting; and resistance to bacterial disease,fungal disease or viral disease; (b) selecting one or more progenyplants that have the desired trait; (c) backcrossing the selectedprogeny plants with a ‘Elidia’ plant to produce backcross progenyplants; (d) selecting for backcross progeny plants that have the desiredtrait and all of the physiological and morphological characteristics ofbasil variety ‘Elidia’; and (e) repeating steps (c) and (d) two or moretimes in succession to produce selected third or higher backcrossprogeny plants that comprise the desired trait. Certain embodiments arealso directed to basil plants produced by such methods, where the plantshave the desired trait and all of the physiological and morphologicalcharacteristics of basil variety In certain embodiments, the desiredtrait is herbicide resistance and the resistance is conferred to anherbicide selected from glyphosate, sulfonylurea, imidazolinone,dicamba, glufosinate, phenoxy proprionic acid, L-phosphinothricin,cyclohexone, cyclohexanedione, triazine, and benzonitrile. In otherembodiments, the desired trait is insect or pest resistance and theinsect or pest resistance is conferred by a transgene encoding aBacillus thuringiensis endotoxin.

In another embodiment, the present invention provides for single geneconverted plants of ‘Elidia’. The single transferred gene may preferablybe a dominant or recessive allele. Preferably, the single transferredgene will confer such traits as male sterility, herbicide resistance,insect or pest resistance, modified fatty acid metabolism, modifiedcarbohydrate metabolism, resistance for bacterial, fungal, or viraldisease, male fertility, enhanced nutritional quality, and industrialusage. The single gene may be a naturally occurring basil gene or atransgene introduced through genetic engineering techniques.

In a further embodiment, the present invention relates to methods fordeveloping basil plants in a basil plant breeding program using plantbreeding techniques including recurrent selection, backcrossing,pedigree breeding, restriction fragment length polymorphism enhancedselection, genetic marker enhanced selection, and transformation. Seeds,basil plants, and parts thereof, produced by such breeding methods arealso part of the invention.

In another embodiment, the present invention is directed to basil,Ocimum basilicum, seed designated as ‘Keira’. In one embodiment, thepresent invention is directed to an Ocimum basilicum basil plant andparts isolated therefrom produced by growing ‘Keira’ basil seed. Inanother embodiment, the present invention is directed to an Ocimumbasilicum plant and parts isolated therefrom having all thephysiological and morphological characteristics of a Lactuca sativaplant produced by growing ‘Keira’ basil seed. In still anotherembodiment, the present invention is directed to an F₁ hybrid Ocimumbasilicum basil seed, plants grown from the seed, and a head isolatedtherefrom having ‘Keira’ as a parent, where ‘Keira’ is grown from‘Keira’ basil seed.

Basil plant parts include basil stems, basil leaves, parts of basilleaves, pollen, ovules, flowers, roots, cells, and the like. In anotherembodiment, the present invention is further directed to basil stems,basil leaves, parts of basil leaves, flowers, pollen, and ovules, roots,or cells isolated from ‘Keira’ basil plants. In another embodiment, thepresent invention is further directed to tissue culture of ‘Keira’ basilplants, and to basil plants regenerated from the tissue culture, wherethe plant has all of the morphological and physiological characteristicsof ‘Keira’ basil plants.

In still another embodiment, the present invention is further directedto packaging material containing ‘Keira’ plant parts. Such packagingmaterial includes but is not limited to boxes, plastic bags, etc. The‘Keira’ plant parts may be combined with other plant parts of otherplant varieties. In yet another embodiment, the present invention isfurther directed to a method of selecting basil plants, by a) growing‘Keira’ basil plants where the ‘Keira’ plants are grown from basil seedand b) selecting a plant from step a). In another embodiment, thepresent invention is further directed to basil plants, plant parts andseeds produced by the basil plants where the basil plants are isolatedby the selection method of the invention. In another embodiment, thepresent invention is further directed to a method of breeding basilplants by crossing a basil plant with a plant grown from ‘Keira’ basilseed. In still another embodiment, the present invention is furtherdirected to basil plants, basil parts from the basil plants, and seedsproduced therefrom where the basil plant is isolated by the breedingmethod of the invention. In some embodiments, the basil plant isolatedby the breeding method is a transgenic basil plant. In anotherembodiment, the present invention is directed to methods for producing abasil plant containing in its genetic material one or more transgenesand to the transgenic basil plant produced by those methods. In anotherembodiment, the present invention is directed to methods for producing amale sterile basil plant by introducing a nucleic acid molecule thatconfers male sterility into a basil plant produced by growing ‘Keira’basil seed, and to male sterile basil plants produced by such methods.

In another embodiment, the present invention is directed to methods ofproducing an herbicide resistant basil plant by introducing a geneconferring herbicide resistance into a basil plant produced by growing‘Keira’ basil seed, where the gene is selected from glyphosate,sulfonylurea, imidazolinone, dicamba, glufosinate, phenoxy proprionicacid, L-phosphinothricin, cyclohexone, cyclohexanedione, triazine, andbenzonitrile. Certain embodiments are also directed to herbicideresistant basil plants produced by such methods. In another embodiment,the present invention is directed to methods of producing a pest orinsect resistant basil plan by introducing a gene conferring pest orinsect resistance into a basil plant produced by growing ‘Keira’ basilseed, and to pest or insect resistant basil plants produced by suchmethods. In certain embodiments, the gene conferring pest or insectresistance encodes a Bacillus thuringiensis endotoxin. In anotherembodiment, the present invention is directed to methods of producing adisease resistant basil plant by introducing a gene conferring diseaseresistance into a basil plant produced by growing ‘Keira’ basil seed,and to disease resistant basil plants produced by such methods. Inanother embodiment, the present invention is directed to methods ofproducing a basil plant with a value-added trait by introducing a geneconferring a value-added trait into a basil plant produced by growing‘Keira’ basil seed, where the gene encodes a protein selected from aferritin, a nitrate reductase, and a monellin. Certain embodiments arealso directed to basil plants having a value-added trait produced bysuch methods.

In another embodiment, the present invention is directed to methods ofintroducing a desired trait into basil variety ‘Keira’, by: (a) crossinga ‘Keira’ plant, with a plant of another basil variety that contains adesired trait to produce progeny plants, where the desired trait isselected from male sterility; herbicide resistance; insect or pestresistance; modified bolting; and resistance to bacterial disease,fungal disease or viral disease; (b) selecting one or more progenyplants that have the desired trait; (c) backcrossing the selectedprogeny plants with a ‘Keira’ plant to produce backcross progeny plants;(d) selecting for backcross progeny plants that have the desired traitand all of the physiological and morphological characteristics of basilvariety ‘Keira’; and (e) repeating steps (c) and (d) two or more timesin succession to produce selected third or higher backcross progenyplants that comprise the desired trait. Certain embodiments are alsodirected to basil plants produced by such methods, where the plants havethe desired trait and all of the physiological and morphologicalcharacteristics of basil variety ‘Keira’. In certain embodiments, thedesired trait is herbicide resistance and the resistance is conferred toan herbicide selected from glyphosate, sulfonylurea, imidazolinone,dicamba, glufosinate, phenoxy proprionic acid, L-phosphinothricin,cyclohexone, cyclohexanedione, triazine, and benzonitrile. In otherembodiments, the desired trait is insect or pest resistance and theinsect or pest resistance is conferred by a transgene encoding aBacillus thuringiensis endotoxin.

In another embodiment, the present invention provides for single geneconverted plants of ‘Keira’. The single transferred gene may preferablybe a dominant or recessive allele. Preferably, the single transferredgene will confer such traits as male sterility, herbicide resistance,insect or pest resistance, modified fatty acid metabolism, modifiedcarbohydrate metabolism, resistance for bacterial, fungal, or viraldisease, male fertility, enhanced nutritional quality, and industrialusage. The single gene may be a naturally occurring basil gene or atransgene introduced through genetic engineering techniques.

In a further embodiment, the present invention relates to methods fordeveloping basil plants in a basil plant breeding program using plantbreeding techniques including recurrent selection, backcrossing,pedigree breeding, restriction fragment length polymorphism enhancedselection, genetic marker enhanced selection, and transformation. Seeds,basil plants, and parts thereof, produced by such breeding methods arealso part of the invention.

In another embodiment, the present invention is directed to basil,Ocimum basilicum, seed designated as ‘Eleonora’, representative sampleof seed having been deposited under ATCC Accession Number PTA-122087. Inone embodiment, the present invention is directed to an Ocimum basilicumbasil plant and parts isolated therefrom produced by growing ‘Eleonora’basil seed. In another embodiment, the present invention is directed toan Ocimum basilicum plant and parts isolated therefrom having all thephysiological and morphological characteristics of a Lactuca sativaplant produced by growing ‘Eleonora’ basil seed having ATCC AccessionNumber PTA-122087. In still another embodiment, the present invention isdirected to an F₁ hybrid Ocimum basilicum basil seed, plants grown fromthe seed, and a head isolated therefrom having ‘Eleonora’ as a parent,where ‘Eleonora’ is grown from ‘Eleonora’ basil seed having ATCCAccession Number PTA-122087.

Basil plant parts include basil stems, basil leaves, parts of basilleaves, pollen, ovules, flowers, roots, cells, and the like. In anotherembodiment, the present invention is further directed to basil stems,basil leaves, parts of basil leaves, flowers, pollen, and ovules, roots,or cells isolated from ‘Eleonora’ basil plants. In another embodiment,the present invention is further directed to tissue culture of‘Eleonora’ basil plants, and to basil plants regenerated from the tissueculture, where the plant has all of the morphological and physiologicalcharacteristics of ‘Eleonora’ basil plants.

In still another embodiment, the present invention is further directedto packaging material containing ‘Eleonora’ plant parts. Such packagingmaterial includes but is not limited to boxes, plastic bags, etc. The‘Eleonora’ plant parts may be combined with other plant parts of otherplant varieties. In yet another embodiment, the present invention isfurther directed to a method of selecting basil plants, by a) growing‘Eleonora’ basil plants where the ‘Eleonora’ plants are grown from basilseed having ATCC Accession Number PTA-122087 and b) selecting a plantfrom step a). In another embodiment, the present invention is furtherdirected to basil plants, plant parts and seeds produced by the basilplants where the basil plants are isolated by the selection method ofthe invention. In another embodiment, the present invention is furtherdirected to a method of breeding basil plants by crossing a basil plantwith a plant grown from ‘Eleonora’ basil seed having ATCC AccessionNumber PTA-122087. In still another embodiment, the present invention isfurther directed to basil plants, basil parts from the basil plants, andseeds produced therefrom where the basil plant is isolated by thebreeding method of the invention. In some embodiments, the basil plantisolated by the breeding method is a transgenic basil plant. In anotherembodiment, the present invention is directed to methods for producing abasil plant containing in its genetic material one or more transgenesand to the transgenic basil plant produced by those methods. In anotherembodiment, the present invention is directed to methods for producing amale sterile basil plant by introducing a nucleic acid molecule thatconfers male sterility into a basil plant produced by growing ‘Eleonora’basil seed, and to male sterile basil plants produced by such methods.

In another embodiment, the present invention is directed to methods ofproducing an herbicide resistant basil plant by introducing a geneconferring herbicide resistance into a basil plant produced by growing‘Eleonora’ basil seed, where the gene is selected from glyphosate,sulfonylurea, imidazolinone, dicamba, glufosinate, phenoxy proprionicacid, L-phosphinothricin, cyclohexone, cyclohexanedione, triazine, andbenzonitrile. Certain embodiments are also directed to herbicideresistant basil plants produced by such methods. In another embodiment,the present invention is directed to methods of producing a pest orinsect resistant basil plan by introducing a gene conferring pest orinsect resistance into a basil plant produced by growing ‘Eleonora’basil seed, and to pest or insect resistant basil plants produced bysuch methods. In certain embodiments, the gene conferring pest or insectresistance encodes a Bacillus thuringiensis endotoxin. In anotherembodiment, the present invention is directed to methods of producing adisease resistant basil plant by introducing a gene conferring diseaseresistance into a basil plant produced by growing ‘Eleonora’ basil seed,and to disease resistant basil plants produced by such methods. Inanother embodiment, the present invention is directed to methods ofproducing a basil plant with a value-added trait by introducing a geneconferring a value-added trait into a basil plant produced by growing‘Eleonora’ basil seed, where the gene encodes a protein selected from aferritin, a nitrate reductase, and a monellin. Certain embodiments arealso directed to basil plants having a value-added trait produced bysuch methods.

In another embodiment, the present invention is directed to methods ofintroducing a desired trait into basil variety ‘Eleonora’, by: (a)crossing a ‘Eleonora’ plant, where a sample of ‘Eleonora’ basil seed wasdeposited under ATCC Accession Number PTA-122087, with a plant ofanother basil variety that contains a desired trait to produce progenyplants, where the desired trait is selected from male sterility;herbicide resistance; insect or pest resistance; modified bolting; andresistance to bacterial disease, fungal disease or viral disease; (b)selecting one or more progeny plants that have the desired trait; (c)backcrossing the selected progeny plants with a ‘Eleonora’ plant toproduce backcross progeny plants; (d) selecting for backcross progenyplants that have the desired trait and all of the physiological andmorphological characteristics of basil variety ‘Eleonora’; and (e)repeating steps (c) and (d) two or more times in succession to produceselected third or higher backcross progeny plants that comprise thedesired trait. Certain embodiments are also directed to basil plantsproduced by such methods, where the plants have the desired trait andall of the physiological and morphological characteristics of basilvariety ‘Eleonora’. In certain embodiments, the desired trait isherbicide resistance and the resistance is conferred to an herbicideselected from glyphosate, sulfonylurea, imidazolinone, dicamba,glufosinate, phenoxy proprionic acid, L-phosphinothricin, cyclohexone,cyclohexanedione, triazine, and benzonitrile. In other embodiments, thedesired trait is insect or pest resistance and the insect or pestresistance is conferred by a transgene encoding a Bacillus thuringiensisendotoxin.

In another embodiment, the present invention provides for single geneconverted plants of ‘Eleonora’. The single transferred gene maypreferably be a dominant or recessive allele. Preferably, the singletransferred gene will confer such traits as male sterility, herbicideresistance, insect or pest resistance, modified fatty acid metabolism,modified carbohydrate metabolism, resistance for bacterial, fungal, orviral disease, male fertility, enhanced nutritional quality, andindustrial usage. The single gene may be a naturally occurring basilgene or a transgene introduced through genetic engineering techniques.

In a further embodiment, the present invention relates to methods fordeveloping basil plants in a basil plant breeding program using plantbreeding techniques including recurrent selection, backcrossing,pedigree breeding, restriction fragment length polymorphism enhancedselection, genetic marker enhanced selection, and transformation. Seeds,basil plants, and parts thereof, produced by such breeding methods arealso part of the invention.

In addition to the exemplary aspects and embodiments described above,further aspects and embodiments will become apparent by reference bystudy of the following description.

DETAILED DESCRIPTION OF THE INVENTION

There are numerous steps in the development of novel, desirable basilgermplasm. Plant breeding begins with the analysis of problems andweaknesses of current basil germplasms, the establishment of programgoals, and the definition of specific breeding objectives. The next stepis selection of germplasm that possess the traits to meet the programgoals. The goal is to combine in a single variety or hybrid an improvedcombination of desirable traits from the parental germplasm. Theseimportant traits may include increased head size and weight, higher seedyield, improved color, resistance to diseases and insects, tolerance todrought and heat, and better agronomic quality.

Choice of breeding or selection methods can depend on the mode of plantreproduction, the heritability of the trait(s) being improved, and thetype of variety used commercially (e.g., F₁ hybrid variety, purelinevariety, etc.). For highly heritable traits, a choice of superiorindividual plants evaluated at a single location will be effective,whereas for traits with low heritability, selection should be based onmean values obtained from replicated evaluations of families of relatedplants. Popular selection methods commonly include pedigree selection,modified pedigree selection, mass selection, and recurrent selection.

The complexity of inheritance influences choice of the breeding method.Backcross breeding is used to transfer one or a few favorable genes fora highly heritable trait into a desirable variety. This approach hasbeen used extensively for breeding disease-resistant varieties. Variousrecurrent selection techniques are used to improve quantitativelyinherited traits controlled by numerous genes. The use of recurrentselection in self-pollinating crops depends on the ease of pollination,the frequency of successful hybrids from each pollination, and thenumber of hybrid offspring from each successful cross.

Each breeding program may include a periodic, objective evaluation ofthe efficiency of the breeding procedure. Evaluation criteria varydepending on the goal and objectives, and can include gain fromselection per year based on comparisons to an appropriate standard, theoverall value of the advanced breeding lines, and the number ofsuccessful varieties produced per unit of input (e.g., per year, perdollar expended, etc.).

Promising advanced breeding lines may be thoroughly tested and comparedto appropriate standards in environments representative of thecommercial target area(s) for at least three years. The best lines canthen be candidates for new commercial varieties. Those still deficientin a few traits may be used as parents to produce new populations forfurther selection. These processes, which lead to the final step ofmarketing and distribution, may take from ten to twenty years from thetime the first cross or selection is made.

One goal of basil plant breeding is to develop new, unique, andgenetically superior basil varieties. A breeder can initially select andcrosses two or more parental lines, followed by repeated selfing andselection, producing many new genetic combinations. Moreover, a breedercan generate multiple different genetic combinations by crossing,selfing, and mutations. A plant breeder can then select which germplasmsto advance to the next generation. These germplasms may then be grownunder different geographical, climatic, and soil conditions, and furtherselections can be made during, and at the end of, the growing season.

The development of commercial basil varieties thus requires thedevelopment of parental basil varieties, the crossing of thesevarieties, and the evaluation of the crosses. Pedigree breeding andrecurrent selection breeding methods may be used to develop varietiesfrom breeding populations. Breeding programs can be used to combinedesirable traits from two or more varieties or various broad-basedsources into breeding pools from which new varieties are developed byselfing and selection of desired phenotypes. The new varieties arecrossed with other varieties and the hybrids from these crosses areevaluated to determine which have commercial potential.

Pedigree breeding is generally used for the improvement ofself-pollinating crops or inbred lines of cross-pollinating crops. Twoparents which possess favorable, complementary traits are crossed toproduce an F₁. An F₂ population is produced by selfing one or severalF₁'s or by intercrossing two F₁'s (sib mating). Selection of the bestindividuals is usually begun in the F₂ population. Then, beginning inthe F₃, the best individuals in the best families are selected.Replicated testing of families, or hybrid combinations involvingindividuals of these families, often follows in the F₄ generation toimprove the effectiveness of selection for traits with low heritability.At an advanced stage of inbreeding (i.e., F₆ and F₇), the best lines ormixtures of phenotypically similar lines are tested for potentialrelease as new varieties.

Mass and recurrent selections can be used to improve populations ofeither self- or cross-pollinating crops. A genetically variablepopulation of heterozygous individuals is either identified or createdby intercrossing several different parents. The best plants are selectedbased on individual superiority, outstanding progeny, or excellentcombining ability. The selected plants are intercrossed to produce a newpopulation in which further cycles of selection are continued.

Backcross breeding may be used to transfer genes for a simply inherited,highly heritable trait into a desirable homozygous cultivar or line thatis the recurrent parent. The source of the trait to be transferred iscalled the donor parent. The resulting plant is expected to have theattributes of the recurrent parent (e.g., cultivar) and the desirabletrait transferred from the donor parent. After the initial cross,individuals possessing the phenotype of the donor parent are selectedand repeatedly crossed (backcrossed) to the recurrent parent. Theresulting plant is expected to have the attributes of the recurrentparent (e.g., cultivar) and the desirable trait transferred from thedonor parent.

The single-seed descent procedure in the strict sense refers to plantinga segregating population, harvesting a sample of one seed per plant, andusing the one-seed sample to plant the next generation. When thepopulation has been advanced from the F₂ to the desired level ofinbreeding, the plants from which lines are derived will each trace todifferent F₂ individuals. The number of plants in a population declineswith each generation due to failure of some seeds to germinate or someplants to produce at least one seed. As a result, not all of the F₂plants originally sampled in the population will be represented by aprogeny when generation advance is completed.

In addition to phenotypic observations, the genotype of a plant can alsobe examined. There are many laboratory-based techniques known in the artthat are available for the analysis, comparison and characterization ofplant genotype. Such techniques include, without limitation, IsozymeElectrophoresis, Restriction Fragment Length Polymorphisms (RFLPs),Randomly Amplified Polymorphic DNAs (RAPDs), Arbitrarily PrimedPolymerase Chain Reaction (AP-PCR), DNA Amplification Fingerprinting(DAF), Sequence Characterized Amplified Regions (SCARs), AmplifiedFragment Length polymorphisms (AFLPs), Simple Sequence Repeats (SSRs,which are also referred to as Microsatellites), and Single NucleotidePolymorphisms (SNPs).

Molecular markers can also be used during the breeding process for theselection of qualitative traits. For example, markers closely linked toalleles or markers containing sequences within the actual alleles ofinterest can be used to select plants that contain the alleles ofinterest during a backcrossing breeding program. The markers can also beused to select toward the genome of the recurrent parent and against themarkers of the donor parent. This procedure attempts to minimize theamount of genome from the donor parent that remains in the selectedplants. It can also be used to reduce the number of crosses back to therecurrent parent needed in a backcrossing program. The use of molecularmarkers in the selection process is often called genetic marker enhancedselection or marker-assisted selection. Molecular markers may also beused to identify and exclude certain sources of germplasm as parentalvarieties or ancestors of a plant by providing a means of trackinggenetic profiles through crosses.

Mutation breeding may also be used to introduce new traits into basilvarieties. Mutations that occur spontaneously or are artificiallyinduced can be useful sources of variability for a plant breeder. Thegoal of artificial mutagenesis is to increase the rate of mutation for adesired characteristic. Mutation rates can be increased by manydifferent means including temperature, long-term seed storage, tissueculture conditions, radiation (such as X-rays, Gamma rays, neutrons,Beta radiation, or ultraviolet radiation), chemical mutagens (such asbase analogs like 5-bromo-uracil), antibiotics, alkylating agents (suchas sulfur mustards, nitrogen mustards, epoxides, ethyleneamines,sulfates, sulfonates, sulfones, or lactones), azide, hydroxylamine,nitrous acid, or acridines. Once a desired trait is observed throughmutagenesis the trait may then be incorporated into existing germplasmby traditional breeding techniques. Details of mutation breeding can befound in Principles of Cultivar Development by Fehr, MacmillanPublishing Company (1993).

The production of double haploids can also be used for the developmentof homozygous varieties in a breeding program. Double haploids areproduced by the doubling of a set of chromosomes from a heterozygousplant to produce a completely homozygous individual. For example, seeWan, et al., Theor. Appl. Genet., 77:889-892 (1989).

Additional non-limiting examples of breeding methods that may be usedinclude, without limitation, those found in Principles of PlantBreeding, John Wiley and Son, pp. 115-161 (1960); Allard (1960);Simmonds (1979); Sneep, et al. (1979); Fehr (1987); and “Carrots andRelated Vegetable Umbelliferae,” Rubatzky, V. E., et al. (1999).

Definitions

In the description that follows, a number of terms are used. In order toprovide a clear and consistent understanding of the specification andclaims, including the scope to be given such terms, the followingdefinitions are provided:

Allele. The allele is any of one or more alternative forms of a gene,all of which relate to one trait or characteristic. In a diploid cell ororganism, the two alleles of a given gene occupy corresponding loci on apair of homologous chromosomes.

Backcrossing. Backcrossing is a process in which a breeder repeatedlycrosses hybrid progeny back to one of the parents, for example, a firstgeneration hybrid F₁ with one of the parental genotype of the F₁ hybrid.

Bolting. The premature development of a flowering stalk, and subsequentseed, before a plant produces a food crop. Bolting is typically causedby late planting when temperatures are low enough to cause vernalizationof the plants.

Bremia lactucae. An oomycete that causes downy mildew in leafyvegetables, such as lettuce and basil, in cooler growing regions.

Cotyledon. One of the first leaves of the embryo of a seed plant;typically one or more in monocotyledons, two in dicotyledons, and two ormore in gymnosperms.

Essentially all the physiological and morphological characteristics. Aplant having essentially all the physiological and morphologicalcharacteristics means a plant having the physiological and morphologicalcharacteristics of the recurrent parent, except for the characteristicsderived from the converted gene.

First water date. The date the seed first receives adequate moisture togerminate. This can and often does equal the planting date.

Fusarium. Any of several fungi of the genus, Fusarium, which are causalagents of stem and root rot, such as Fusarium wilt. Fusarium wilt ischaracterized by damping-off, collapse of the plant, wilting, and abrown dry rot.

Gene. As used herein, “gene” refers to a segment of nucleic acid. A genecan be introduced into a genome of a species, whether from a differentspecies or from the same species, using transformation or variousbreeding methods.

Maturity date. Maturity refers to the stage when the plants are of fullsize or optimum weight, in marketable form or shape to be of commercialor economic value.

Quantitative Trait Loci. Quantitative Trait Loci (QTL) refers to geneticloci that control to some degree, numerically representable traits thatare usually continuously distributed.

Regeneration. Regeneration refers to the development of a plant fromtissue culture.

RHS. RHS refers to the Royal Horticultural Society of England whichpublishes an official botanical color chart quantitatively identifyingcolors according to a defined numbering system. The chart may bepurchased from Royal Horticulture Society Enterprise Ltd., RHS Garden;Wisley, Woking; Surrey GU236QB, UK.

Single gene converted. Single gene converted or conversion plant refersto plants which are developed by a plant breeding technique calledbackcrossing or via genetic engineering where essentially all of thedesired morphological and physiological characteristics of a line arerecovered in addition to the single gene transferred into the line viathe backcrossing technique or via genetic engineering.

Overview of the Variety ‘Elidia’

Basil variety ‘Elidia’ is a basil variety suitable for greenhouse, openfield, and tunnel production. It is the result of numerous generationsof plant selections chosen for its yield, medium leaf blade glossiness,and intermediate resistance to Fusarium (Fusarium oxysporum f. sp.Basilicum).

The variety has shown uniformity and stability for the traits, withinthe limits of environmental influence for the traits. It has beenself-pollinated a sufficient number of generations with carefulattention to uniformity of plant type. The line has been increased withcontinued observation for uniformity. No variant traits have beenobserved or are expected in variety ‘Elidia’.

Objective Description of the Variety ‘Elidia’

Basil variety ‘Elidia’ has the following morphologic and othercharacteristics:

Plant:

-   -   Habit: Erect; similar to varieties ‘Genovese’, ‘Grand vert’, and        ‘Zöldgömb’    -   Total height: Medium; similar to variety ‘Lemon’    -   Density: Medium; similar to varieties ‘Lemon’ and        ‘Keskenylevelü’

Stem:

-   -   Anthocyanin coloration: Absent; similar to variety ‘Grand vert’    -   Hairiness: Absent; similar to variety ‘A feuille de laitue’    -   Number of flowering shoots (at full flowering): More than three;        similar to variety ‘True Thai’

Leaves:

-   -   Blade shape: Ovate; similar to variety ‘Fin vert’    -   Blade length: Long; similar to variety ‘Géant Mammouth’    -   Blade width: Medium; similar to variety ‘Genovese’    -   Blade anthocyanin coloration of upper side: Absent; similar to        varieties ‘Grand vert’ and ‘Zöldgömb’    -   Blade green color: Medium; similar to varieties ‘Fin vert nain’        and ‘Lemon’    -   Blade glossiness: Medium; similar to variety ‘Osmin’    -   Blade blistering: Medium; similar to varieties ‘Genovese’ and        ‘Grand vert’    -   Blade profile in cross section: Convex; similar to varieties        ‘Genovese’ and ‘Grand vert’    -   Blade serration of margin: Present; similar to variety ‘Purple        Ruffles’    -   Blade depth of serration: Shallow; similar to ‘Italian Large        Leaf’    -   Blade undulation of margin: Absent or very weak; similar to        variety ‘Grand vert’

Petioles:

-   -   Length: Medium; similar to variety ‘Genovese’

Flowering Stems:

-   -   Average length of internodes (at end of flowering): Medium;        similar to variety ‘Grand vert’    -   Total length (at end of flowering): Medium; similar to variety        ‘Genovese’    -   Hairiness of bracts: Present; similar to variety ‘Lemon’

Flowers:

-   -   Color of corolla: White; similar to varieties ‘Genovese’ and        ‘Grand vert’    -   Color of style: White; similar to variety ‘Genovese’    -   Time of flowering (10% of plants flowering): Medium; similar to        varieties ‘Genovese’ and ‘Grand vert’

Disease/Pest Resistance:

-   -   Fusarium (Fusarium oxysporum f sp. Basilicum): Intermediate        resistance        Comparison of ‘Elidia’ to Commercial Basil Variety

Table 1 below compares some of the characteristics of basil variety‘Elidia’ with the basil variety ‘Eowyn’. Column 1 lists thecharacteristic, column 2 shows the characteristics for basil variety‘Elidia’, and column 3 shows the characteristics for basil variety‘Eowyn’.

TABLE 1 Characteristic ‘Elidia’ ‘Eowyn’ Leaf blade glossiness MediumStrongOverview of the Variety ‘Keira’

Basil variety ‘Keira’ is a basil variety suitable for greenhouse, openfield, and tunnel production. It is the result of numerous generationsof plant selections chosen for its yield, medium leaf blade blistering,medium leaf blade glossiness, and ability to grow well under coolercultivation conditions.

The variety has shown uniformity and stability for the traits, withinthe limits of environmental influence for the traits. It has beenself-pollinated a sufficient number of generations with carefulattention to uniformity of plant type. The line has been increased withcontinued observation for uniformity. No variant traits have beenobserved or are expected in variety ‘Keira’.

Objective Description of the Variety ‘Keira’

Basil variety ‘Keira’ has the following morphologic and othercharacteristics:

Plant:

-   -   Habit: Erect; similar to varieties ‘Genovese’, ‘Grand vert’, and        ‘Zöldgömb’    -   Total height: Tall; similar to varieties ‘Genovese’ and ‘Grand        vert’    -   Density: Medium; similar to varieties ‘Lemon’ and        ‘Keskenylevelü’

Stem:

-   -   Anthocyanin coloration: Absent; similar to variety ‘Grand vert’    -   Hairiness: Absent; similar to variety ‘A feuille de laitue’    -   Number of flowering shoots (at full flowering): More than three;        similar to variety ‘True Thai’

Leaves:

-   -   Blade shape: Ovate; similar to variety ‘Fin vert’    -   Blade length: Medium; similar to variety ‘Osmin’    -   Blade width: Medium; similar to variety ‘Genovese’    -   Blade anthocyanin coloration of upper side: Absent; similar to        varieties ‘Grand vert’ and ‘Zöldgömb’    -   Blade green color: Dark; similar to variety ‘Sweet Thai’    -   Blade glossiness: Medium; similar to variety ‘Osmin’    -   Blade blistering: Medium; similar to varieties ‘Genovese’ and        ‘Grand vert’    -   Blade profile in cross section: Convex; similar to varieties        ‘Genovese’ and ‘Grand vert’    -   Blade serration of margin: Present; similar to variety ‘Purple        Ruffles’    -   Blade depth of serration: Shallow; similar to ‘Italian Large        Leaf’    -   Blade undulation of margin: Absent or very weak; similar to        variety ‘Grand vert’

Petioles:

-   -   Length: Medium; similar to variety ‘Genovese’

Flowering Stems:

-   -   Average length of internodes (at end of flowering): Medium;        similar to variety ‘Grand vert’    -   Total length (at end of flowering): Medium; similar to variety        ‘Genovese’    -   Hairiness of bracts: Present; similar to variety ‘Lemon’

Flowers:

-   -   Color of corolla: White; similar to varieties ‘Genovese’ and        ‘Grand vert’    -   Color of style: White; similar to variety ‘Genovese’    -   Time of flowering (10% of plants flowering): Medium; similar to        varieties ‘Genovese’ and ‘Grand vert’        Comparison of ‘Keira’ to Commercial Basil Variety

Table 2 below compares some of the characteristics of basil variety‘Keira’ with the basil variety ‘Eowyn’. Column 1 lists thecharacteristics, column 2 shows the characteristics for basil variety‘Keira’, and column 3 shows the characteristics for basil variety‘Eowyn’.

TABLE 2 Characteristic ‘Keira’ ‘Eowyn’ Leaf blade blistering MediumStrong Leaf blade glossiness Medium StrongOverview of the Variety ‘Eleonora’

Basil variety ‘Eleonora’ is a basil variety suitable for greenhouse,open field, and tunnel production. It is the result of numerousgenerations of plant selections chosen for its yield, medium leaf bladegreen color, weak leaf blade blistering, and intermediate resistance todowny mildew (Peronospora belbahrii).

The variety has shown uniformity and stability for the traits, withinthe limits of environmental influence for the traits. It has beenself-pollinated a sufficient number of generations with carefulattention to uniformity of plant type. The line has been increased withcontinued observation for uniformity. No variant traits have beenobserved or are expected in variety ‘Eleonora’.

Objective Description of the Variety ‘Eleonora’

Basil variety ‘Eleonora’ has the following morphologic and othercharacteristics:

Plant:

-   -   Habit: Erect; similar to varieties ‘Genovese’, ‘Grand vert’, and        ‘Zöldgömb’    -   Total height: Tall; similar to varieties ‘Genovese’ and ‘Grand        vert’    -   Density: Medium; similar to varieties ‘Lemon’ and        ‘Keskenylevelü’

Stem:

-   -   Anthocyanin coloration: Absent; similar to variety ‘Grand vert’    -   Hairiness: Absent; similar to variety ‘A feuille de laitue’    -   Number of flowering shoots (at full flowering): More than three;        similar to variety ‘True Thai’

Leaves:

-   -   Blade shape: Ovate; similar to variety ‘Fin vert’    -   Blade length: Medium; similar to variety ‘Osmin’    -   Blade width: Medium; similar to variety ‘Genovese’    -   Blade anthocyanin coloration of upper side: Absent; similar to        varieties ‘Grand vert’ and ‘Zöldgömb’    -   Blade green color: Medium; similar to varieties ‘Fin vert nain’        and ‘Lemon’    -   Blade glossiness: Medium; similar to variety ‘Osmin’    -   Blade blistering: Weak; similar to varieties ‘Dark Opal’ and        ‘Keskenylevelü’    -   Blade profile in cross section: Convex; similar to varieties        ‘Genovese’ and ‘Grand vert’    -   Blade serration of margin: Present; similar to variety ‘Purple        Ruffles’    -   Blade depth of serration: Shallow; similar to ‘Italian Large        Leaf’    -   Blade undulation of margin: Absent or very weak; similar to        variety ‘Grand vert’

Petioles:

-   -   Length: Medium; similar to variety ‘Genovese’

Flowering Stems:

-   -   Average length of internodes (at end of flowering): Medium;        similar to variety ‘Grand vert’    -   Total length (at end of flowering): Medium; similar to variety        ‘Genovese’    -   Hairiness of bracts: Present; similar to variety ‘Lemon’

Flowers:

-   -   Color of corolla: White; similar to varieties ‘Genovese’ and        ‘Grand vert’    -   Color of style: White; similar to variety ‘Genovese’    -   Time of flowering (10% of plants flowering): Early; similar to        variety ‘Keskenylevelü’

Disease/Pest Resistance:

-   -   Downy Mildew (Peronospora belbahrii): Intermediate resistance        Comparison of ‘Eleonora’ to Commercial Basil Variety

Table 3 below compares some of the characteristics of basil variety‘Eleonora’ with the basil variety ‘Edwina’. Column 1 lists thecharacteristics, column 2 shows the characteristics for basil variety‘Eleonora’, and column 3 shows the characteristics for basil variety‘Edwina’.

TABLE 3 Characteristic ‘Eleonora’ ‘Edwina’ Leaf blade green color MediumDark Leaf blade blistering Weak MediumFurther Embodiments

Additional methods include, but are not limited to, expression vectorsintroduced into plant tissues using a direct gene transfer method, suchas microprojectile-mediated delivery, DNA injection, electroporation,and the like. More preferably, expression vectors are introduced intoplant tissues by using either microprojectile-mediated delivery with abiolistic device or by using Agrobacterium-mediated transformation.Transformed plants obtained with the protoplasm of the invention areintended to be within the scope of this invention.

With the advent of molecular biological techniques that have allowed theisolation and characterization of genes that encode specific proteinproducts, scientists in the field of plant biology developed a stronginterest in engineering the genome of plants to contain and expressforeign genes, or additional, or modified versions of native, orendogenous, genes (perhaps driven by different promoters) in order toalter the traits of a plant in a specific manner. Any DNA sequences,whether from a different species or from the same species, which areintroduced into the genome using transformation or various breedingmethods, are referred to herein collectively as “transgenes.” Over thelast fifteen to twenty years, several methods for producing transgenicplants have been developed, and the present invention, in particularembodiments, also relates to transformed versions of the claimed line.

Nucleic acids or polynucleotides refer to RNA or DNA that is linear orbranched, single or double stranded, or a hybrid thereof. The term alsoencompasses RNA/DNA hybrids. These terms also encompass untranslatedsequence located at both the 3′ and 5′ ends of the coding region of thegene: at least about 1000 nucleotides of sequence upstream from the 5′end of the coding region and at least about 200 nucleotides of sequencedownstream from the 3′ end of the coding region of the gene. Less commonbases, such as inosine, 5-methylcytosine, 6-methyladenine, hypoxanthine,and others can also be used for antisense, dsRNA, and ribozyme pairing.For example, polynucleotides that contain C-5 propyne analogues ofuridine and cytidine have been shown to bind RNA with high affinity andto be potent antisense inhibitors of gene expression. Othermodifications, such as modification to the phosphodiester backbone, orthe 2′-hydroxy in the ribose sugar group of the RNA can also be made.The antisense polynucleotides and ribozymes can consist entirely ofribonucleotides, or can contain mixed ribonucleotides anddeoxyribonucleotides. The polynucleotides of the invention may beproduced by any means, including genomic preparations, cDNApreparations, in vitro synthesis, RT-PCR, and in vitro or in vivotranscription.

Plant transformation involves the construction of an expression vectorthat will function in plant cells. Such a vector contains DNA thatcontains a gene under control of, or operatively linked to, a regulatoryelement (for example, a promoter). The expression vector may contain oneor more such operably linked gene/regulatory element combinations. Thevector(s) may be in the form of a plasmid, and can be used alone or incombination with other plasmids, to provide transformed basil plantsusing transformation methods as described below to incorporatetransgenes into the genetic material of the basil plant(s).

Expression Vectors for Basil Transformation: Marker Genes

Expression vectors include at least one genetic marker, operably linkedto a regulatory element (for example, a promoter) that allowstransformed cells containing the marker to be either recovered bynegative selection, i.e., inhibiting growth of cells that do not containthe selectable marker gene, or by positive selection, i.e., screeningfor the product encoded by the genetic marker. Many commonly usedselectable marker genes for plant transformation are well known in thetransformation arts, and include, for example, genes that code forenzymes that metabolically detoxify a selective chemical agent which maybe an antibiotic or an herbicide, or genes that encode an altered targetwhich is insensitive to the inhibitor. A few positive selection methodsare also known in the art.

One commonly used selectable marker gene for plant transformation is theneomycin phosphotransferase II (nptII) gene, isolated from transposonTn5, which when placed under the control of plant regulatory signalsconfers resistance to kanamycin. Fraley, et al., PNAS, 80:4803 (1983).Another commonly used selectable marker gene is the hygromycinphosphotransferase gene which confers resistance to the antibiotichygromycin. Vanden Elzen, et al., Plant Mol. Biol., 5:299 (1985).

Additional selectable marker genes of bacterial origin that conferresistance to antibiotics include gentamycin acetyl transferase,streptomycin phosphotransferase, aminoglycoside-3′-adenyl transferase,the bleomycin resistance determinant. Hayford, et al., Plant Physiol.,86:1216 (1988); Jones, et al., Mol. Gen. Genet., 210:86 (1987); Svab, etal., Plant Mol. Biol., 14:197 (1990); Hille, et al., Plant Mol. Biol.,7:171 (1986). Other selectable marker genes confer resistance toherbicides such as glyphosate, glufosinate, or bromoxynil. Comai, etal., Nature, 317:741-744 (1985); Gordon-Kamm, et al., Plant Cell,2:603-618 (1990); and Stalker, et al., Science, 242:419-423 (1988).

Selectable marker genes for plant transformation that are not ofbacterial origin include, for example, mouse dihydrofolate reductase,plant 5-enolpyruvylshikimate-3-phosphate synthase, and plantacetolactate synthase. Eichholtz, et al., Somatic Cell Mol. Genet.,13:67 (1987); Shah, et al., Science, 233:478 (1986); and Charest, etal., Plant Cell Rep., 8:643 (1990).

Another class of marker genes for plant transformation requiresscreening of presumptively transformed plant cells rather than directgenetic selection of transformed cells for resistance to a toxicsubstance such as an antibiotic. These genes are particularly useful toquantify or visualize the spatial pattern of expression of a gene inspecific tissues and are frequently referred to as reporter genesbecause they can be fused to a gene or gene regulatory sequence for theinvestigation of gene expression. Commonly used genes for screeningpresumptively transformed cells include α-glucuronidase (GUS),α-galactosidase, luciferase, chloramphenicol, and acetyltransferase.Jefferson, R. A., Plant Mol. Biol., 5:387 (1987); Teeri, et al., EMBOJ., 8:343 (1989); Koncz, et al., PNAS, 84:131 (1987); and DeBlock, etal., EMBO J., 3:1681 (1984).

In vivo methods for visualizing GUS activity that do not requiredestruction of plant tissues are available. Molecular Probes,Publication 2908, IMAGENE GREEN, pp. 1-4 (1993) and Naleway, et al., J.Cell Biol., 115:151a (1991). However, these in vivo methods forvisualizing GUS activity have not proven useful for recovery oftransformed cells because of low sensitivity, high fluorescentbackgrounds, and limitations associated with the use of GUS genes asselectable markers.

More recently, a gene encoding Green Fluorescent Protein (GFP) has beenutilized as a marker for gene expression in prokaryotic and eukaryoticcells. Chalfie, et al., Science, 263:802 (1994). GFP and mutants of GFPmay be used as screenable markers.

Expression Vectors for Basil Transformation: Promoters

Genes included in expression vectors must be driven by a nucleotidesequence containing a regulatory element (for example, a promoter).Several types of promoters are now well known in the transformationarts, as are other regulatory elements that can be used alone or incombination with promoters.

As used herein, “promoter” includes reference to a region of DNAupstream from the start of transcription and involved in recognition andbinding of RNA polymerase and other proteins to initiate transcription.A “plant promoter” is a promoter capable of initiating transcription inplant cells. Examples of promoters under developmental control includepromoters that preferentially initiate transcription in certain tissues,such as leaves, roots, seeds, fibers, xylem vessels, tracheids, orsclerenchyma. Such promoters are referred to as “tissue-preferred,”Promoters which initiate transcription only in certain tissue arereferred to as “tissue-specific.” A “cell type” specific promoterprimarily drives expression in certain cell types in one or more organs,for example, vascular cells in roots or leaves. An “inducible” promoteris a promoter which is under environmental control. Examples ofenvironmental conditions that may affect transcription by induciblepromoters include anaerobic conditions or the presence of light.Tissue-specific, tissue-preferred, cell type specific, and induciblepromoters constitute the class of “non-constitutive” promoters. A“constitutive” promoter is a promoter which is active under mostenvironmental conditions.

A. Inducible Promoters:

An inducible promoter is operably linked to a gene for expression inbasil. Optionally, the inducible promoter is operably linked to anucleotide sequence encoding a signal sequence which is operably linkedto a gene for expression in basil. With an inducible promoter, the rateof transcription increases in response to an inducing agent.

Any inducible promoter can be used in the instant invention. See Ward,et al., Plant Mol. Biol., 22:361-366 (1993). Exemplary induciblepromoters include, but are not limited to, that from the ACEI systemwhich responds to copper (Meft, et al., PNAS, 90:4567-4571 (1993)); In2gene from maize which responds to benzenesulfonamide herbicide safeners(Hershey, et al., Mol. Gen. Genet., 227:229-237 (1991) and Gatz, et al.,Mol. Gen. Genet., 243:32-38 (1994)) or Tet repressor from Tn10 (Gatz, etal., Mol. Gen. Genet., 227:229-237 (1991)). A particularly preferredinducible promoter is a promoter that responds to an inducing agent towhich plants do not normally respond. An exemplary inducible promoter isthe inducible promoter from a steroid hormone gene, the transcriptionalactivity of which is induced by a glucocorticosteroid hormone. Schena,et al., PNAS, 88:0421 (1991).

B. Constitutive Promoters:

A constitutive promoter is operably linked to a gene for expression inbasil or the constitutive promoter is operably linked to a nucleotidesequence encoding a signal sequence which is operably linked to a genefor expression in basil.

Many different constitutive promoters can be utilized in the instantinvention. Exemplary constitutive promoters include, but are not limitedto, the promoters from plant viruses such as the 35S promoter from CaMV(Odell, et al., Nature, 313:810-812 (1985)) and the promoters from suchgenes as rice actin (McElroy, et al., Plant Cell, 2:163-171 (1990));ubiquitin (Christensen, et al., Plant Mol. Biol., 12:619-632 (1989) andChristensen, et al., Plant Mol. Biol., 18:675-689 (1992)); pEMU (Last,et al., Theor. Appl. Genet., 81:581-588 (1991)); MAS (Velten, et al.,EMBO 1, 3:2723-2730 (1984)) and maize H3 histone (Lepetit, et al., Mol.Gen. Genet., 231:276-285 (1992) and Atanassova, et al., Plant J., 2(3):291-300 (1992)). The ALS promoter, XbaI/NcoI fragment 5′ to theBrassica napus ALS3 structural gene (or a nucleotide sequence similarityto said XbaI/NcoI fragment), represents a particularly usefulconstitutive promoter. See PCT Application No. WO 96/30530.

C. Tissue-Specific or Tissue-Preferred Promoters:

A tissue-specific promoter is operably linked to a gene for expressionin basil. Optionally, the tissue-specific promoter is operably linked toa nucleotide sequence encoding a signal sequence which is operablylinked to a gene for expression in basil. Plants transformed with a geneof interest operably linked to a tissue-specific promoter produce theprotein product of the transgene exclusively, or preferentially, in aspecific tissue.

Any tissue-specific or tissue-preferred promoter can be utilized in theinstant invention. Exemplary tissue-specific or tissue-preferredpromoters include, but are not limited to, a root-preferred promoter,such as that from the phaseolin gene (Murai, et al., Science, 23:476-482(1983) and Sengupta-Gopalan, et al., PNAS, 82:3320-3324 (1985)); aleaf-specific and light-induced promoter such as that from cab orrubisco (Simpson, et al., EMBO J., 4(11):2723-2729 (1985) and Timko, etal., Nature, 318:579-582 (1985)); an anther-specific promoter such asthat from LAT52 (Twell, et al., Mol. Gen. Genet., 217:240-245 (1989)); apollen-specific promoter such as that from Zm13 (Guerrero, et al., Mol.Gen. Genet., 244:161-168 (1993)) or a microspore-preferred promoter suchas that from apg (Twell, et al., Sex. Plant Reprod., 6:217-224 (1993)).

Signal Sequences for Targeting Proteins to Subcellular Compartments

Transport of protein produced by transgenes to a subcellular compartmentsuch as the chloroplast, vacuole, peroxisome, glyoxysome, cell wall, ormitochondrion, or for secretion into the apoplast, is accomplished bymeans of operably linking the nucleotide sequence encoding a signalsequence to the 5′ and/or 3′ region of a gene encoding the protein ofinterest. Targeting sequences at the 5′ and/or 3′ end of the structuralgene may determine, during protein synthesis and processing, where theencoded protein is ultimately compartmentalized.

The presence of a signal sequence directs a polypeptide to either anintracellular organelle or subcellular compartment or for secretion tothe apoplast. Many signal sequences are known in the art. See, forexample, Becker, et al., Plant Mol. Biol., 20:49 (1992); Close, P. S.,Master's Thesis, Iowa State University (1993); Knox, C., et al.,“Structure and Organization of Two Divergent Alpha-Amylase Genes fromBarley,” Plant Mol. Biol., 9:3-17 (1987); Lerner, et al., PlantPhysiol., 91:124-129 (1989); Fontes, et al., Plant Cell, 3:483-496(1991); Matsuoka, et al., PNAS, 88:834 (1991); Gould, et al., J. Cell.Biol., 108:1657 (1989); Creissen, et al., Plant 2:129 (1991); Kalderon,et al., A short amino acid sequence able to specify nuclear location,Cell, 39:499-509 (1984); and Steifel, et al., Expression of a maize cellwall hydroxyproline-rich glycoprotein gene in early leaf and rootvascular differentiation, Plant Cell, 2:785-793 (1990).

Foreign Protein Genes and Agronomic Genes

With transgenic plants according to the present invention, a foreignprotein can be produced in commercial quantities. Thus, techniques forthe selection and propagation of transformed plants, which are wellunderstood in the art, yield a plurality of transgenic plants which areharvested in a conventional manner, and a foreign protein then can beextracted from a tissue of interest or from total biomass. Proteinextraction from plant biomass can be accomplished by known methods whichare discussed, for example, by Heney and Orr, Anal. Biochem., 114:92-6(1981).

According to a preferred embodiment, the transgenic plant provided forcommercial production of foreign protein is basil. In another preferredembodiment, the biomass of interest is seed. For the relatively smallnumber of transgenic plants that show higher levels of expression, agenetic map can be generated, primarily via conventional RFLP, PCR, andSSR analysis, which identifies the approximate chromosomal location ofthe integrated DNA molecule. For exemplary methodologies in this regard,see Methods in Plant Molecular Biology and Biotechnology, Glick andThompson Eds., 269:284, CRC Press, Boca Raton (1993). Map informationconcerning chromosomal location is useful for proprietary protection ofa subject transgenic plant. If unauthorized propagation is undertakenand crosses made with other germplasm, the map of the integration regioncan be compared to similar maps for suspect plants, to determine if thelatter have a common parentage with the subject plant. Map comparisonswould involve hybridizations, RFLP, PCR, SSR, and sequencing, all ofwhich are conventional techniques.

Likewise, by means of the present invention, agronomic genes can beexpressed in transformed plants. More particularly, plants can begenetically engineered to express various phenotypes of agronomicinterest. Exemplary genes implicated in this regard include, but are notlimited to, those categorized below:

A. Genes that Confer Resistance to Pests or Disease and that Encode:

1. Plant disease resistance genes. Plant defenses are often activated byspecific interaction between the product of a disease resistance gene(R) in the plant and the product of a corresponding avirulence (Avr)gene in the pathogen. A plant line can be transformed with a clonedresistance gene to engineer plants that are resistant to specificpathogen strains. See, for example, Jones, et al., Science, 266:789(1994) (cloning of the tomato Cf-9 gene for resistance to Cladosporiumfulvum); Martin, et al., Science, 262:1432 (1993) (tomato Pto gene forresistance to Pseudomonas syringae pv. tomato encodes a protein kinase);and Mindrinos, et al., Cell, 78:1089 (1994) (Arabidopsis RSP2 gene forresistance to Pseudomonas syringae).

2. A Bacillus thuringiensis protein, a derivative thereof, or asynthetic polypeptide modeled thereon. See, for example, Geiser, et al.,Gene, 48:109 (1986), who disclose the cloning and nucleotide sequence ofa Bt δ-endotoxin gene. Moreover, DNA molecules encoding δ-endotoxingenes can be purchased from American Type Culture Collection, Manassas,Va., for example, under ATCC Accession Nos. 40098, 67136, 31995, and31998.

3. A lectin. See, for example, the disclosure by Van Damme, et al.,Plant Mol. Biol., 24:25 (1994), who disclose the nucleotide sequences ofseveral Clivia miniata mannose-binding lectin genes.

4. A vitamin-binding protein such as avidin. See PCT Application No. US93/06487, the contents of which are hereby incorporated by reference.The application teaches the use of avidin and avidin homologues aslarvicides against insect pests.

5. An enzyme inhibitor, for example, a protease or proteinase inhibitor,or an amylase inhibitor. See, for example, Abe, et al., J Biol. Chem.,262:16793 (1987) (nucleotide sequence of rice cysteine proteinaseinhibitor); Huub, et al., Plant Mol. Biol., 21:985 (1993) (nucleotidesequence of cDNA encoding tobacco proteinase inhibitor I); and Sumitani,et al., Biosci. Biotech. Biochem., 57:1243 (1993) (nucleotide sequenceof Streptomyces nitrosporeus α-amylase inhibitor).

6. An insect-specific hormone or pheromone, such as an ecdysteroid andjuvenile hormone, a variant thereof, a mimetic based thereon, or anantagonist or agonist thereof. See, for example, the disclosure byHammock, et al., Nature, 344:458 (1990), of baculovirus expression ofcloned juvenile hormone esterase, an inactivator of juvenile hormone.

7. An insect-specific peptide or neuropeptide which, upon expression,disrupts the physiology of the affected pest. For example, see thedisclosures of Regan, J. Biol. Chem., 269:9 (1994) (expression cloningyields DNA coding for insect diuretic hormone receptor) and Pratt, etal., Biochem. Biophys. Res. Comm., 163:1243 (1989) (an allostatin isidentified in Diploptera puntata). See also, U.S. Pat. No. 5,266,317 toTomalski, et al., who disclose genes encoding insect-specific, paralyticneurotoxins.

8. An insect-specific venom produced in nature, by a snake, a wasp, etc.For example, see Pang, et al., Gene, 116:165 (1992), for disclosure ofheterologous expression in plants of a gene coding for a scorpioninsectotoxic peptide.

9. An enzyme responsible for a hyper-accumulation of a monoterpene, asesquiterpene, a steroid, hydroxamic acid, a phenylpropanoid derivative,or another non-protein molecule with insecticidal activity.

10. An enzyme involved in the modification, including thepost-translational modification, of a biologically active molecule; forexample, a glycolytic enzyme, a proteolytic enzyme, a lipolytic enzyme,a nuclease, a cyclase, a transaminase, an esterase, a hydrolase, aphosphatase, a kinase, a phosphorylase, a polymerase, an elastase, achitinase, and a glucanase, whether natural or synthetic. See PCTApplication No. WO 93/02197 in the name of Scott, et al., whichdiscloses the nucleotide sequence of a callase gene. DNA molecules whichcontain chitinase-encoding sequences can be obtained, for example, fromthe ATCC under Accession Nos. 39637 and 67152. See also, Kramer, et al.,Insect Biochem. Mol. Biol., 23:691 (1993), who teach the nucleotidesequence of a cDNA encoding tobacco hornworm chitinase, and Kawalleck,et al., Plant Mol. Biol., 21:673 (1993), who provide the nucleotidesequence of the lettuce ubi4-2 polyubiquitin gene.

11. A molecule that stimulates signal transduction. For example, see thedisclosure by Botella, et al., Plant Mol. Biol., 24:757 (1994), ofnucleotide sequences for mung bean calmodulin cDNA clones, and Griess,et al., Plant Physiol., 104:1467 (1994), who provide the nucleotidesequence of a maize calmodulin cDNA clone.

12. A hydrophobic moment peptide. See PCT Application No. WO 95/16776(disclosure of peptide derivatives of tachyplesin which inhibit fungalplant pathogens) and PCT Application No. WO 95/18855 (teaches syntheticantimicrobial peptides that confer disease resistance), the respectivecontents of which are hereby incorporated by reference.

13. A membrane permease, a channel former, or a channel blocker. Forexample, see the disclosure of Jaynes, et al., Plant Sci., 89:43 (1993),of heterologous expression of a cecropin-β, lytic peptide analog torender transgenic tobacco plants resistant to Pseudomonas solanacearum.

14. A viral-invasive protein or a complex toxin derived therefrom. Forexample, the accumulation of viral coat proteins in transformed plantcells imparts resistance to viral infection and/or disease developmenteffected by the virus from which the coat protein gene is derived, aswell as by related viruses. See Beachy, et al., Ann. Rev. Phytopathol.,28:451 (1990). Coat protein-mediated resistance has been conferred upontransformed plants against alfalfa mosaic virus, cucumber mosaic virus,tobacco streak virus, potato virus X, potato virus Y, tobacco etchvirus, tobacco rattle virus, and tobacco mosaic virus. Id.

15. An insect-specific antibody or an immunotoxin derived therefrom.Thus, an antibody targeted to a critical metabolic function in theinsect gut would inactivate an affected enzyme, killing the insect. SeeTaylor, et al., Abstract #497, Seventh Int'l Symposium on MolecularPlant-Microbe Interactions, Edinburgh, Scotland (1994) (enzymaticinactivation in transgenic tobacco via production of single-chainantibody fragments).

16. A virus-specific antibody. See, for example, Tavladoraki, et al.,Nature, 366:469 (1993), who show that transgenic plants expressingrecombinant antibody genes are protected from virus attack.

17. A developmental-arrestive protein produced in nature by a pathogenor a parasite. Thus, fungal endo-α-1, 4-D-polygalacturonases facilitatefungal colonization and plant nutrient released by solubilizing plantcell wall homo-α-1,4-D-galacturonase. See Lamb, et al., Bio/technology,10:1436 (1992). The cloning and characterization of a gene which encodesa bean endopolygalacturonase-inhibiting protein is described by Toubart,et al., Plant J, 2:367 (1992).

18. A developmental-arrestive protein produced in nature by a plant. Forexample, Logemann, et al., Bio/technology, 10:305 (1992), have shownthat transgenic plants expressing the barley ribosome-inactivating genehave an increased resistance to fungal disease.

Any of the above listed disease or pest resistance genes (1-19) can beintroduced into the claimed basil variety through a variety of meansincluding but not limited to transformation and crossing.

B. Genes that Confer Resistance to an Herbicide:

1. An herbicide that inhibits the growing point or meristem, such as animidazolinone or a sulfonylurea. Exemplary genes in this category codefor mutant ALS and AHAS enzyme as described, for example, by Lee, etal., EMBO J., 7:1241 (1988) and Miki, et al., Theor. Appl. Genet.,80:449 (1990), respectively.

2. Glyphosate (resistance conferred by mutant5-enolpyruvlshikimate-3-phosphate synthase (EPSPS) and aroA genes,respectively) and other phosphono compounds, such as glufosinate(phosphinothricin acetyl transferase (PAT), dicamba and Streptomyceshygroscopicus phosphinothricin-acetyl transferase PAT bar genes), andpyridinoxy or phenoxy proprionic acids and cyclohexones (ACCaseinhibitor-encoding genes). See, for example, U.S. Pat. No. 4,940,835 toShah, et al., which discloses the nucleotide sequence of a form of EPSPSwhich can confer glyphosate resistance. A DNA molecule encoding a mutantaroA gene can be obtained under ATCC Accession No. 39256, and thenucleotide sequence of the mutant gene is disclosed in U.S. Pat. No.4,769,061 to Comai. See also, Umaballava-Mobapathie in TransgenicResearch, 8:1, 33-44 (1999) that discloses Lactuca sativa resistant toglufosinate. European Patent Application No. 0 333 033 to Kumada, etal., and U.S. Pat. No. 4,975,374 to Goodman, et al., disclose nucleotidesequences of glutamine synthetase genes which confer resistance toherbicides, such as L-phosphinothricin. The nucleotide sequence of aphosphinothricin-acetyl-transferase gene is provided in EuropeanApplication No. 0 242 246 to Leemans, et al. DeGreef, et al.,Bio/technology, 7:61 (1989), describe the production of transgenicplants that express chimeric bar genes coding for phosphinothricinacetyl transferase activity. Exemplary of genes conferring resistance tophenoxy proprionic acids and cyclohexones, such as sethoxydim andhaloxyfop, are the Acc1-S1, Acc1-S2, and Acc1-S3 genes described byMarshall, et al., Theor. Appl. Genet., 83:435 (1992).

3. An herbicide that inhibits photosynthesis, such as a triazine (psbAand gs+ genes) and a benzonitrile (nitrilase gene). Przibilla, et al.,Plant Cell, 3:169 (1991), describe the transformation of Chlamydomonaswith plasmids encoding mutant psbA genes. Nucleotide sequences fornitrilase genes are disclosed in U.S. Pat. No. 4,810,648 to Stalker, andDNA molecules containing these genes are available under ATCC AccessionNos. 53435, 67441, and 67442. Cloning and expression of DNA coding for aglutathione S-transferase is described by Hayes, et al., Biochem. J.,285:173 (1992).

4. Acetohydroxy acid synthase, which has been found to make plants thatexpress this enzyme resistant to multiple types of herbicides, has beenintroduced into a variety of plants. See Hattori, et al., Mol. Gen.Genet., 246:419 (1995). Other genes that confer tolerance to herbicidesinclude a gene encoding a chimeric protein of rat cytochrome P4507A1 andyeast NADPH-cytochrome P450 oxidoreductase (Shiota, et al., PlantPhysiol., 106:17 (1994)), genes for glutathione reductase and superoxidedismutase (Aono, et al., Plant Cell Physiol., 36:1687 (1995)), and genesfor various phosphotransferases (Datta, et al., Plant Mol. Biol., 20:619(1992)).

5. Protoporphyrinogen oxidase (protox) is necessary for the productionof chlorophyll, which is necessary for all plant survival. The protoxenzyme serves as the target for a variety of herbicidal compounds. Theseherbicides also inhibit growth of all the different species of plantspresent, causing their total destruction. The development of plantscontaining altered protox activity which are resistant to theseherbicides are described in U.S. Pat. Nos. 6,288,306, 6,282,837,5,767,373, and International Publication WO 01/12825.

Any of the above listed herbicide genes (1-5) can be introduced into theclaimed basil variety through a variety of means including, but notlimited to, transformation and crossing.

C. Genes that Confer or Contribute to a Value-Added Trait, Such as:

1. Increased iron content of the basil, for example, by introducing intoa plant a soybean ferritin gene as described in Goto, et al., ActaHorticulturae., 521, 101-109 (2000).

2. Decreased nitrate content of leaves, for example, by introducing intoa basil plant a gene coding for a nitrate reductase. See, for example,Curtis, et al., Plant Cell Rep., 18:11, 889-896 (1999).

3. Increased sweetness of the basil by introducing a gene coding formonellin that elicits a flavor 100,000 times sweeter than sugar on amolar basis. See Penarrubia, et al., Bio/technology, 10:561-564 (1992).

4. Modified fatty acid metabolism, for example, by introducing into aplant an antisense gene of stearyl-ACP desaturase to increase stearicacid content of the plant. See Knultzon, et al., PNAS, 89:2625 (1992).

5. Modified carbohydrate composition effected, for example, byintroducing into plants a gene coding for an enzyme that alters thebranching pattern of starch. See Shiroza, et al., J. Bacteriol., 170:810(1988) (nucleotide sequence of Streptococcus mutantsfructosyltransferase gene); Steinmetz, et al., Mol. Gen. Genet., 20:220(1985) (nucleotide sequence of Bacillus subtilis levansucrase gene);Pen, et al., Bio/technology, 10:292 (1992) (production of transgenicplants that express Bacillus lichenifonnis α-amylase); Elliot, et al.,Plant Mol. Biol., 21:515 (1993) (nucleotide sequences of tomatoinvertase genes); Søgaard, et al., J Biol. Chem., 268:22480 (1993)(site-directed mutagenesis of barley α-amylase gene); and Fisher, etal., Plant Physiol., 102:1045 (1993) (maize endosperm starch branchingenzyme II).

D. Genes that Control Male-Sterility:

1. Introduction of a deacetylase gene under the control of atapetum-specific promoter and with the application of the chemicalN-Ac-PPT. See International Publication WO 01/29237.

2. Introduction of various stamen-specific promoters. See InternationalPublications WO 92/13956 and WO 92/13957.

3. Introduction of the barnase and the barstar genes. See Paul, et al.,Plant Mol. Biol., 19:611-622 (1992).

Methods for Basil Transformation

Numerous methods for plant transformation have been developed, includingbiological and physical, plant transformation protocols. See, forexample, Mild, et al., “Procedures for Introducing Foreign DNA intoPlants” in Methods in Plant Molecular Biology and Biotechnology, Glickand Thompson Eds., CRC Press, Inc., Boca Raton, pp. 67-88 (1993). Inaddition, expression vectors and in vitro culture methods for plant cellor tissue transformation and regeneration of plants are available. See,for example, Gruber, et al., “Vectors for Plant Transformation” inMethods in Plant Molecular Biology and Biotechnology, Glick and ThompsonEds., CRC Press, Inc., Boca Raton, pp. 89-119 (1993).

A. Agrobacterium-Mediated Transformation:

One method for introducing an expression vector into plants is based onthe natural transformation system of Agrobacterium. See, for example,Horsch, et al., Science, 227:1229 (1985); Curtis, et al., Journal ofExperimental Botany, 45:279, 1441-1449 (1994); Torres, et al., PlantCell Tissue and Organ Culture, 34:3, 279-285 (1993); and Dinant, et al.,Molecular Breeding, 3:1, 75-86 (1997). A. tumefaciens and A. rhizogenesare plant pathogenic soil bacteria which genetically transform plantcells. The Ti and Ri plasmids of A. tumefaciens and A. rhizogenes,respectively, carry genes responsible for genetic transformation of theplant. See, for example, Kado, C. I., Crit. Rev. Plant Sci., 10:1(1991). Descriptions of Agrobacterium vector systems and methods forAgrobacterium-mediated gene transfer are provided by Gruber, et al.,supra, Miki, et al., supra, and Moloney, et al., Plant Cell Rep., 8:238(1989). See also, U.S. Pat. No. 5,591,616 issued Jan. 7, 1997.

B. Direct Gene Transfer:

Several methods of plant transformation collectively referred to asdirect gene transfer have been developed as an alternative toAgrobacterium-mediated transformation. A generally applicable method ofplant transformation is microprojectile-mediated transformation whereDNA is carried on the surface of microprojectiles measuring 1 μm to 4μm. The expression vector is introduced into plant tissues with abiolistic device that accelerates the microprojectiles to speeds of 300m/s to 600 m/s which is sufficient to penetrate plant cell walls andmembranes. Russell, D. R., et al., Plant Cell Rep., 12 (3, January),165-169 (1993); Aragao, F. J. L., et al., Plant Mol. Biol., 20 (2,October), 357-359 (1992); Aragao, F. J. L., et al., Plant Cell Rep., 12(9, July), 483-490 (1993); Aragao, Theor. Appl. Genet., 93:142-150(1996); Kim, J., Minamikawa, T., Plant Sci., 117:131-138 (1996);Sanford, et al., Part. Sci. Technol., 5:27 (1987); Sanford, J. C.,Trends Biotech., 6:299 (1988); Klein, et al., Bio/technology, 6:559-563(1988); Sanford, J. C., Physiol. Plant, 7:206 (1990); Klein, et al.,Bio/technology, 10:268 (1992).

Another method for physical delivery of DNA to plants is sonication oftarget cells. Zhang, et al., Bio/technology, 9:996 (1991).Alternatively, liposome and spheroplast fusion have been used tointroduce expression vectors into plants. Deshayes, et al., EMBO J.,4:2731 (1985) and Christou, et al., PNAS, 84:3962 (1987). Direct uptakeof DNA into protoplasts using CaCl₂ precipitation, polyvinyl alcohol, orpoly-L-ornithine has also been reported. Hain, et al., Mol. Gen. Genet.,199:161 (1985) and Draper, et al., Plant Cell Physiol., 23:451 (1982).Electroporation of protoplasts and whole cells and tissues have alsobeen described. Saker, M., Kuhne, T., Biologia Plantarum, 40(4):507-514(1997/98); Donn, et al., In Abstracts of VIIth International Congress onPlant Cell and Tissue Culture IAPTC, A2-38, p. 53 (1990); D'Halluin, etal., Plant Cell, 4:1495-1505 (1992); and Spencer, et al., Plant Mol.Biol., 24:51-61 (1994). See also Chupean, et al., Bio/technology, 7:5,503-508 (1989).

Following transformation of basil target tissues, expression of theabove-described selectable marker genes allows for preferentialselection of transformed cells, tissues and/or plants, usingregeneration and selection methods now well known in the art.

The foregoing methods for transformation would typically be used forproducing a transgenic line. The transgenic line could then be crossedwith another (non-transformed or transformed) line in order to produce anew transgenic basil line. Alternatively, a genetic trait which has beenengineered into a particular basil variety using the foregoingtransformation techniques could be introduced into another line usingtraditional backcrossing techniques that are well known in the plantbreeding arts. For example, a backcrossing approach could be used tomove an engineered trait from a public, non-elite inbred line into anelite inbred line, or from an inbred line containing a foreign gene inits genome into an inbred line or lines which do not contain that gene.As used herein, “crossing” can refer to a simple X by Y cross, or theprocess of backcrossing, depending on the context.

Gene Conversions

When the term “basil plant” is used in the context of the presentinvention, this also includes any gene conversions of that variety. Theterm “gene converted plant” as used herein refers to those basil plantswhich are developed by backcrossing, genetic engineering, or mutation,where essentially all of the desired morphological and physiologicalcharacteristics of a variety are recovered in addition to the one ormore genes transferred into the variety via the backcrossing technique,genetic engineering, or mutation. Backcrossing methods can be used withthe present invention to improve or introduce a characteristic into thevariety. The term “backcrossing” as used herein refers to the repeatedcrossing of a hybrid progeny back to the recurrent parent, i.e.,backcrossing 1, 2, 3, 4, 5, 6, 7, 8, 9, or more times to the recurrentparent. The parental basil plant which contributes the gene for thedesired characteristic is termed the “nonrecurrent” or “donor parent.”This terminology refers to the fact that the nonrecurrent parent is usedone time in the backcross protocol and therefore does not recur. Theparental basil plant to which the gene or genes from the nonrecurrentparent are transferred is known as the recurrent parent as it is usedfor several rounds in the backcrossing protocol. Poehlman & Sleper(1994) and Fehr (1993). In a typical backcross protocol, the originalvariety of interest (recurrent parent) is crossed to a second variety(nonrecurrent parent) that carries the gene of interest to betransferred. The resulting progeny from this cross are then crossedagain to the recurrent parent and the process is repeated until a basilplant is obtained where essentially all of the desired morphological andphysiological characteristics of the recurrent parent are recovered inthe converted plant, in addition to the transferred gene from thenonrecurrent parent.

The selection of a suitable recurrent parent is an important step for asuccessful backcrossing procedure. The goal of a backcross protocol isto alter or substitute a trait or characteristic in the original line.To accomplish this, a gene of the recurrent variety is modified orsubstituted with the desired gene from the nonrecurrent parent, whileretaining essentially all of the rest of the desired genetic, andtherefore the desired physiological and morphological, constitution ofthe original line. The choice of the particular nonrecurrent parent willdepend on the purpose of the backcross. One of the major purposes is toadd some commercially desirable, agronomically important trait to theplant. The exact backcrossing protocol will depend on the characteristicor trait being altered to determine an appropriate testing protocol.Although backcrossing methods are simplified when the characteristicbeing transferred is a dominant allele, a recessive allele may also betransferred. In this instance it may be necessary to introduce a test ofthe progeny to determine if the desired characteristic has beensuccessfully transferred.

Many gene traits have been identified that are not regularly selected inthe development of a new line but that can be improved by backcrossingtechniques. Gene traits may or may not be transgenic. Examples of thesetraits include, but are not limited to, male sterility, modified fattyacid metabolism, modified carbohydrate metabolism, herbicide resistance,resistance for bacterial, fungal, or viral disease, insect resistance,enhanced nutritional quality, industrial usage, yield stability, andyield enhancement. These genes are generally inherited through thenucleus. Several of these gene traits are described in U.S. Pat. Nos.5,777,196, 5,948,957, and 5,969,212, the disclosures of which arespecifically hereby incorporated by reference.

Tissue Culture

Further reproduction of the variety can occur by tissue culture andregeneration. Tissue culture of various tissues of basil andregeneration of plants therefrom is well known and widely published. Forexample, reference may be had to Teng, et al., HortScience, 27:9,1030-1032 (1992); Teng, et al., HortScience, 28:6, 669-1671 (1993);Zhang, et al., Journal of Genetics and Breeding, 46:3, 287-290 (1992);Webb, et al., Plant Cell Tissue and Organ Culture, 38:1, 77-79 (1994);Curtis, et al., Journal of Experimental Botany, 45:279, 1441-1449(1994); Nagata, et al., Journal for the American Society forHorticultural Science, 125:6, 669-672 (2000); and Ibrahim, et al., PlantCell Tissue and Organ Culture, 28(2), 139-145 (1992). It is clear fromthe literature that the state of the art is such that these methods ofobtaining plants are routinely used and have a very high rate ofsuccess. Thus, another aspect of this invention is to provide cellswhich upon growth and differentiation produce basil plants having thephysiological and morphological characteristics of varieties ‘Elidia’,‘Keira’, and ‘Eleonora’.

As used herein, the term “tissue culture” indicates a compositioncontaining isolated cells of the same or a different type or acollection of such cells organized into parts of a plant. Exemplarytypes of tissue cultures are protoplasts, calli, meristematic cells, andplant cells that can generate tissue culture that are intact in plantsor parts of plants, such as leaves, pollen, embryos, roots, root tips,anthers, pistils, flowers, seeds, petioles, suckers, and the like. Meansfor preparing and maintaining plant tissue culture are well known in theart. By way of example, a tissue culture containing organs has been usedto produce regenerated plants. U.S. Pat. Nos. 5,959,185, 5,973,234, and5,977,445 describe certain techniques, the disclosures of which areincorporated herein by reference.

Additional Breeding Methods

The invention is also directed to methods for producing a basil plant bycrossing a first parent basil plant with a second parent basil plantwhere the first or second parent basil plant is a basil plant of variety‘Elidia’, ‘Keira’, or ‘Eleonora’. Further, both first and second parentbasil plants can come from basil variety ‘Elidia’, ‘Keira’, or‘Eleonora’. Thus, any such methods using basil variety ‘Elidia’,‘Keira’, or ‘Eleonora’ are part of the invention: selfing, backcrosses,hybrid production, crosses to populations, and the like. All plantsproduced using basil variety ‘Elidia’, ‘Keira’, or ‘Eleonora’ as atleast one parent are within the scope of this invention, including thosedeveloped from varieties derived from basil variety ‘Elidia’, ‘Keira’,or ‘Eleonora’. Advantageously, this basil variety could be used incrosses with other, different, basil plants to produce the firstgeneration (F₁) basil hybrid seeds and plants with superiorcharacteristics. The variety of the invention can also be used fortransformation where exogenous genes are introduced and expressed by thevariety of the invention. Genetic variants created either throughtraditional breeding methods using basil variety ‘Keira’, or ‘Eleonora’or through transformation of variety ‘Keira’, or ‘Eleonora’ by any of anumber of protocols known to those of skill in the art are intended tobe within the scope of this invention.

The following describes breeding methods that may be used with basilvariety ‘Keira’, or ‘Eleonora’ in the development of further basilplants. One such embodiment is a method for developing variety ‘Keira’,or ‘Eleonora’ progeny basil plants in a basil plant breeding program,by: obtaining the basil plant, or a part thereof, of variety ‘Keira’, or‘Eleonora’, utilizing said plant or plant part as a source of breedingmaterial, and selecting a basil variety ‘Keira’, or ‘Eleonora’ progenyplant with molecular markers in common with variety ‘Keira’, or‘Eleonora’ and/or with morphological and/or physiologicalcharacteristics selected from the characteristics listed in the sectionentitled “Objective description of the variety ‘Elidia’,” “Objectivedescription of the variety ‘Keira’”, or “Objective description of thevariety ‘Eleonora’”. Breeding steps that may be used in the basil plantbreeding program include pedigree breeding, backcrossing, mutationbreeding, and recurrent selection. In conjunction with these steps,techniques such as RFLP-enhanced selection, genetic marker enhancedselection (for example, SSR markers), and the making of double haploidsmay be utilized.

Another method involves producing a population of basil variety ‘Keira’,or ‘Eleonora’ progeny basil plants, by crossing variety ‘Keira’, or‘Eleonora’ with another basil plant, thereby producing a population ofbasil plants, which, on average, derive 50% of their alleles from basilvariety ‘Keira’, or ‘Eleonora’. A plant of this population may beselected and repeatedly selfed or sibbed with a basil variety resultingfrom these successive filial generations. One embodiment of thisinvention is the basil variety produced by this method and that hasobtained at least 50% of its alleles from basil variety ‘Keira’, or‘Eleonora’. One of ordinary skill in the art of plant breeding wouldknow how to evaluate the traits of two plant varieties to determine ifthere is no significant difference between the two traits expressed bythose varieties. For example, see Fehr and Walt, Principles of VarietyDevelopment, pp. 261-286 (1987). Thus the invention includes basilvariety ‘Elidia’, ‘Keira’, or ‘Eleonora’ progeny basil plants containinga combination of at least two variety ‘Keira’, or ‘Eleonora’ traitsselected from those listed in the section entitled “Objectivedescription of the variety ‘Elidia’,” “Objective description of thevariety ‘Keira’”, or “Objective description of the variety ‘Eleonora’”;or the variety ‘Keira’, or ‘Eleonora’ combination of traits listed inthe Summary of the Invention, so that said progeny basil plant is notsignificantly different for said traits than basil variety ‘Keira’, or‘Eleonora’ as determined at the 5% significance level when grown in thesame environmental conditions. Using techniques described herein,molecular markers may be used to identify said progeny plant as a basilvariety ‘Keira’, or ‘Eleonora’ progeny plant. Mean trait values may beused to determine whether trait differences are significant, andpreferably the traits are measured on plants grown under the sameenvironmental conditions. Once such a variety is developed, its value issubstantial since it is important to advance the germplasm base as awhole in order to maintain or improve traits such as yield, diseaseresistance, pest resistance, and plant performance in extremeenvironmental conditions.

Progeny of basil variety ‘Elidia’, ‘Keira’, or ‘Eleonora’ may also becharacterized through their filial relationship with basil variety‘Keira’, or ‘Eleonora’, as for example, being within a certain number ofbreeding crosses of basil variety ‘Keira’, or ‘Eleonora’. A breedingcross is a cross made to introduce new genetics into the progeny, and isdistinguished from a cross, such as a self or a sib cross, made toselect among existing genetic alleles. The lower the number of breedingcrosses in the pedigree, the closer the relationship between basilvariety ‘Keira’, or ‘Eleonora’ and its progeny. For example, progenyproduced by the methods described herein may be within 1, 2, 3, 4, or 5breeding crosses of basil variety ‘Elidia’, ‘Keira’, or ‘Eleonora’.

As used herein, the term “plant” includes plant cells, plantprotoplasts, plant cell tissue cultures from which basil plants can beregenerated, plant calli, plant clumps, and plant cells that are intactin plants or parts of plants, such as leaves, pollen, embryos,cotyledons, hypocotyl, roots, root tips, anthers, pistils, flowers,ovules, seeds, stems, and the like.

The use of the terms “a,” “an,” and “the,” and similar referents in thecontext of describing the invention (especially in the context of thefollowing claims) are to be construed to cover both the singular and theplural, unless otherwise indicated herein or clearly contradicted bycontext. The terms “comprising,” “having,” “including,” and “containing”are to be construed as open-ended terms (i.e., meaning “including, butnot limited to,”) unless otherwise noted. Recitation of ranges of valuesherein are merely intended to serve as a shorthand method of referringindividually to each separate value falling within the range, unlessotherwise indicated herein, and each separate value is incorporated intothe specification as if it were individually recited herein. Forexample, if the range 10-15 is disclosed, then 11, 12, 13, and 14 arealso disclosed. All methods described herein can be performed in anysuitable order unless otherwise indicated herein or otherwise clearlycontradicted by context. The use of any and all examples, or exemplarylanguage (e.g., “such as”) provided herein, is intended merely to betterilluminate the invention and does not pose a limitation on the scope ofthe invention unless otherwise claimed. No language in the specificationshould be construed as indicating any non-claimed element as essentialto the practice of the invention.

While a number of exemplary aspects and embodiments have been discussedabove, those of skill in the art will recognize certain modifications,permutations, additions, and sub-combinations thereof. It is thereforeintended that the following appended claims and claims hereafterintroduced are interpreted to include all such modifications,permutations, additions, and sub-combinations as are within their truespirit and scope.

DEPOSIT INFORMATION

A deposit of the basil variety ‘Eleonora’ is maintained by Enza ZadenUSA, Inc., having an address at 7 Harris Place, Salinas, Calif. 93901,United States. Access to these deposits will be available during thependency of this application to persons determined by the Commissionerof Patents and Trademarks to be entitled thereto under 37 C.F.R. §1.14and 35 U.S.C. §122. Upon allowance of any claims in this application,all restrictions on the availability to the public of variety ‘Eleonora’will be irrevocably removed by affording access to a deposit of at least2,500 seeds of each variety with the American Type Culture Collection,(ATCC), ATCC Patent Depository, 10801 University Boulevard, Manassas,Va., 20110, USA.

Basil Variety ‘Eleonora’

At least 2500 seeds of basil variety ‘Eleonora’ were deposited on Apr.2, 2015 according to the Budapest Treaty in the American Type CultureCollection (ATCC), ATCC Patent Depository, 10801 University Boulevard,Manassas, Va., 20110, USA. The deposit has been assigned ATCC NumberPTA-122087. Access to this deposit will be available during the pendencyof this application to persons determined by the Commissioner of Patentsand Trademarks to be entitled thereto under 37 C.F.R. §1.14 and 35U.S.C. §122. Upon allowance of any claims in this application, allrestrictions on the availability to the public of the variety will beirrevocably removed.

The deposit will be maintained in the ATCC depository, which is a publicdepository, for a period of at least 30 years, or at least 5 years afterthe most recent request for a sample of the deposit, or for theeffective life of the patent, whichever is longer, and will be replacedif a deposit becomes nonviable during that period.

The invention claimed is:
 1. Basil seed designated as ‘Eleonora’,representative sample of seed having been deposited under ATCC AccessionNumber PTA-122087.
 2. A basil plant produced by growing the seed ofclaim
 1. 3. A plant part from the plant of claim
 2. 4. The plant part ofclaim 3 wherein said part is a stem, a leaf, or a portion thereof.
 5. Abasil plant having all the physiological and morphologicalcharacteristics of the basil plant of claim
 2. 6. A plant part from theplant of claim
 5. 7. The plant part of claim 6, wherein said part is astem, a leaf, or a portion thereof.
 8. An F1 hybrid basil plant having‘Eleonora’ as a parent where ‘Eleonora’ is grown from the seed ofclaim
 1. 9. Pollen or an ovule of the plant of claim
 2. 10. A tissueculture of the plant of claim
 2. 11. A basil plant regenerated from thetissue culture of claim 10, wherein the plant has all of themorphological and physiological characteristics of a basil plantproduced by growing seed designated as ‘Eleonora’, representative sampleof seed having been deposited under having ATCC Accession NumberPTA-122087.
 12. A method of making basil seeds, said method comprisingcrossing the plant of claim 2 with another basil plant and harvestingseed therefrom.
 13. A method of making basil variety ‘Eleonora’, saidmethod comprising selecting seeds from the cross of one ‘Eleonora’ plantwith another ‘Eleonora’ plant, a sample of ‘Eleonora’ basil seed havingbeen deposited under ATCC Accession Number PTA-122087.